There is a fundamental gap in understanding how antiviral RIG-like receptors (RLRs) function in innate immunity. Until this gap is filled, it is dificult to precisely and specifically manipulate RLR- based immune pathways to promote health and treat disease. The long-term goal of this research program is to translate detailed knowledge of immune-system function into viable molecular therapies. The objective of this application is to characterize the mechanisms underlying differences in molecular binding and immune responses triggered by RLRs. Based on preliminary data, the central hypothesis is that differences in RLR function have been driven primarily by adaptive modifications of specific contact residues to recognize different viral RNA structures; these changes in the binding properties directly affect the type and intensity of downstream immune response produced. The rationale for the proposed research is that understanding the molecular basis for differences in RLR function will provide opportunities for developing targeted biomedical therapies. The central hypothesis will be tested by pursuing two specific aims: 1) determine how evolution shaped the RNA-binding properties of RLRs, and 2) determine how evolution of RLR- RNA binding affects the cellular immune response. These aims will be achieved using a combination of molecular-functional analyses of extant and ancestral RLRs, characterization of the cellular immune response induced by these RLRs, and evolutionary analysis to understand the molecular basis for RLR functional divergence. This approach is innovative, because it challenges the existing paradigm of developing therapeutic approaches without a complete understanding of how the systems being targeted evolved or function. The proposed research is significant, because it is expected to immediately present opportunities for developing molecular therapies capable of manipulating the cellular immune response with very high specificity and limited off-target effects.